2-amino-4,6-diphenethylamino-s-triazine for treating arteriosclerosis

ABSTRACT

THE PRESENT INVENTION RELATES TO A VALUABLE MEDICAMENT FOR ORAL ADMINISTRATION CONTAINING 2-AMINO-4,6-DIPHENETHYLAMINO-S-TRIAZINE AS AN EFFECTIVE INGREDIENT. THE DRUG OF THIS INVENTION IS USEFUL FOR REDUCING SERUM CHOLESTEROL LEVEL AND FOR PREVENTING ARTERIOSCLEROSIS.

3,816,629 Z-AMINO-4,6-DIPHENETHYLAMINO-S-TRIAZINE FOR TREATING ARTERIOSCLEROSIS Tsutomu Irikura, Kyoichi Higo, Akitoshi Maeda, Fumihiko Morinaga, and Takeshi N aruke, Tokyo, Japan, as-' signors to Kyorin Seiyaku Kabushiki Kaisha, Tokyo, Japan No Drawing. Application Sept. 1, 1972, Ser. No. 285,680,

which is a continuation-in-part of abandoned application Ser. No. 134,394, Apr. 15, 1971. Divided and this application Feb. 12, 1973, Ser. No. 331,861

Int. Cl. A61k 27/00 US. Cl. 424249 1 Claim ABSTRACT OF THE DISCLOSURE The present invention relates to a valuable medicament for oral administration containing 2-amino-4,6-diphenethylamino-s-triazine as an efiective ingredient. The drug of this invention is useful for reducing serum cholesterol level and for preventing arteriosclerosis.

This is a division of application Ser. No. 285,680, filed on Sept. 1, 1972, which in turn is a continuation-in-part of application Ser. No. 134,394, filed on Apr. 15, 1971, now abandoned.

This invention relates to pharmaceutical compositions; more particularly it relates to pharmaceutical compositions which may be of use for the reduction of serum cholesterol and/or the suppression of arteriosclerosis.

The compound, Z-aminno 4,6 diphenethylamino-s-triazine, is described by Hans Suter and Hans Zutter in Helvetica Chimica Acta 48 (8), 1940-1944 (196-5) These authors synthesized twelve s-triazine derivatives expecting to obtain anti-diabetic agents which could be administered orally (derivatives of s-triazine were formed by ring closure of biguanides which had been already known to reduce the blood sugar level). However, as described in the report, they found no effective anti-diabetic agents.

2-Amino-4,6-diphenethylamino-s-triazine was one of the twelve compounds, but no details of its preparation were given, nor was the compound shown to possess any pharmacological activity.

The investigations which lead to the discovery of the present invention were also concerned with s-triazine derivatives, but for reasons quite different from those of Hans Suter and Hans Zutter. Recently, it was found that certain novel derivatives of s-triazine could suppress the abnormally enhanced activity of the reticuloendothelial system of animals induced by bacterial endotoxin. It was also found that there exists a relationship between this effect on hyperfunction of the reticuloendothelial system and the therapeutic efiect of the derivatives on arteriosclerosis. These novel s-triazine derivatives prevent the deposition of lipids on the arterial wall Without reducing the serum cholesterol level. In order that the mechanism of the anti-arterioschlerotic action and the structure activity relationship of s-triazine derivatives could be further investigated.

Other s-triazine derivatives were synthesized and tested to see whether they had any particular actions on experimental arteriosclerosis. This investigation revealed the unexpected fact that some of the s-triazine derivatives shown the ability to reduce the serum cholesterol level. In particular, it was found that 2-amino-4,6-diphenethylamino-striazine suppresses the bio-synthesis of cholesterol in the liver, reduces the serum cholesterol level and prevents the experimental arteriosclerosis.

This compound is the first s-triazine derivative, which has been found to possess such pharmaceutical activity.

P ce

Before sthe compound of this'invention can be usedany possibility-that the nature of the vehicle, considered of course, in relation to the route by which the composition is intended to be administered, could be harmful rather than beneficial.

The choice of a suitable mode of presentation, together with an appropriate vehicle, is believed to be within the competence of those accustomed to the preparation of pharmaceutical formulations.

Accordingly this invention provides a pharmaceutical composition which comprises 2-amino-4,6-diphenethylamino-s-triazine in association with a suitable pharmaceutical vehicle.

The compositions of this invention may be administered orally and in respect of these modes, the pharmaceutical vehicle is preferably: the ingestible excipient of a tablet, coated tablet, sublingual tablet or pill; the ingestible container of a capsule or cachet; the ingestible pulverulent solid carrier of a powder; or the ingestible liquid medium of a syrup, solution, suspension or elixir.

In order further to illustrate this invention but without being limited thereto, the following examples are given:

EXAMPLE 1 Preparation of 2-amino-4,fi-diphenethylamino-s-triazine 3.3 g. of 2-amino-4,6-dichloro-s-triazine were suspended in ml. of water and 9.7 g. of phenethylamine were added dropwise, with stirring, for 2 hours. After cooling the reaction mixture, the precipitate was separated and recrystallized from isopropanol to give 6.0 g. of 2-amino- 4,6-diphenethyla-mino-s-triazine in the form of colourless needles having a melting point of l40-l42 C.

Aanalysis for C H N -Calculated (percent): C, 68.24; H, 6.63; N, 25.13. Determined (percent): C, 68.10; H, 6.54; N, 25.25.

EXAMPLE 2 Tablets are prepared by mixing and granulating in accordance with known pharmaceutical techniques the following ingredients.

Ingredient: Mg./tablet 2-Amino-4,G-diphenethylamino-s-triazine 50 Lactose 7 Crystalline cellulose 25 Potato starch 15 Magnesium stearate 0.5 Hydroxypropyl cellulose 2.5

EXAMPLE 3 Tablets are prepared by mixing and granulating in accordance with known pharmaceutical techniques, for example using the following ingredients.

maceutical techniques from the following ingredients. Ingredient: Mg./capsule 2-Amino-4,6-diphenethylamino-s-triazine 50 Lactose 148 Crystalline cellulose 30 Potato starch 50 Magnesium stearate 2 V N S I I 23 g., were employed. These were divided into 3 groups, Capsules are prepared in accordance with known pharone group normal p) Was fed a basal diet, the maceutical techniques, for example using the following second group (the control group) was fed a cholesterol- 1ngred1ents.- I I v enriched diet (the basal diet with the addition of 2% of In i I M j ul 5 chloresteroland 0.1% of cholic acid), whilst the third 2-Amino-4,6-diphenethylamino s-triazine 60 group was fed the basal diet with the addition'of 2'%of LaC Se V 138 cholesterol, 0.1% of cholic acid and.( 0714% of 2-.amino cl'ystanme cellulose 30 4,6-diphenethylamino-s-triazine. All the. mice were-.fed figg z i g g I g those diets for 14 days and were decapitated on the 14th w x fday. Whole blood was collected and the total chores'te'rol EXAMPLE 6 1 in the serum was determined by the Zak and Henly rrieth One pulverulenta is prepared in accordance with known the results are Shown n Table I. This Table'cle y pharmaceutical techniques from the following ingredients. shows that 2 amino-4,6-diphenethylamino-s-triazine sigl di Mg /Pu1vem1enta 15 nificantly suppresses the increase in the serum cholesterol Z-Amino-4,6-diphenethylamino-s-triazine 50 level in the mice fed a cholesterol-enriched diet.

Lactose 450 i TABLE I Dose or K Increase Food eholes- Total choles- Rate of Number oibody intgke (gmfkeoll 2331;136:2151; sugges: Group g glig a s 15) day) (meaniSnDJ (percent) Nnrmal 9 0.6 160.6 0 17s.3=|=22.5 Control 10 2. 7 189. 7 3. 794 270. 13:31. 9 2-amino 4,6-diphenethylamlno-s-triazine. 8 1.1 163.2 3.264 b215.4:l=30.0 5625 I The rate of suppression was calculated by the following equation:

(Total serum cholesterol of controls) (Total serum cholesterol of mice receiving 2-amino-4,6-

diphenethylamino s-trlazine) Rate of suppression X1 (Total serum cholesterolot controls) (Total serum cholesterol of normals) b Significantly lower than the control group (P 0.005).

EXAMPLE 7 EXPERIMENT B r A suspenslon for oral admmlstratlon is prepared m Effect of 2-amino-4,G-diphenethylamino-s-triazine on theaccordance with known pharmaceutical techniques from 40 serum cholesterol level in normal rats the following mgredlents.

Ingredient: Male Wistar strain rats, 10 weeks old and weighing- 2-Amino-4,G-diphenethylamino-s-tri- 236-300 g., were employed. The tests were conducted azine mg with three groups of rats; these three groups were fed- Sodium carboxymethyl cell lo mg 300 45 diets containing 0.0% (as control), 0.1% and 0.3% of Simple syrup ml 2-amino-4,6-diphenethylamino-s-triazine respectively. The

EXAMPLE 8 rats were all fed for 14 days and decapitated on the 14th Granules are prepared in accordance with known pharas in Experiment A above. The results are shown in Table maceutical techniques from the following ingredients. 50 II.

TABLE II 7 Increase Dose of Weight of liver 1 Oholestero Number of body compound (gm./100 g. in serum Proportion of 6-amino-4,6-diphenethylof rats weight (mg/kg] body weight) (mg. percent) nmino-s-triazine in diet (percent) in group (grn.) day) (meaniS.D.) (mean=l=S.D.)

I I Significantly lowerthan control group (P 0.025).

Ingredient: V Mg. EXPERIMENT C v I {Ziggy henethy1am1194nazme 5 Suppression of the bio-synthesis of choresterol in the liver. :Potam starch v I p 150 of rats by 2-arnino-4,o-diphenethylamino-s-triazine crystallmssellvlose 100 Male Wistar strain rats which were fed for 7 days H 3 }Y Proper quanmy' with diets containing 0.1% and 0.3% respectively of 2- v EXPERIMENT A amino 4,6 diphenethylamino-s-triazine, were compared The eifect of 2-amino 4 6 diphenethylamino s triazine on rats 3 3 2 were z f the g g g wi outtea itionotiscompoun. nte; ay, .g i sfigg a g m mlce fed cholesterol these rats were given 1 ml. per 200 g. body weight of Male" a d-strain mice; 8" weeks old and weighing 20- by injection in the tail vein, and the rats were killed 60 day. Their serum cholesterol levels 'were then determined 1-Cacetate (20 ucurie/ZO ,umole per 1 ml. solution).

minutes later. Their livers were separated and hydrolysed this test. One group (the normal group) was fed a basal at 80 C. with 5 ml. of alcoholic potassium hydiet. droxide solution per 1 gm. of liver for 3 hours. The second group (the control group) was fed a choles- The solutwn obtamed after the hydrolysls was a terol-enriched diet (the basal diet with the addition of rated, and the cholesterol was extracted from the residue 5 2% of cholesterol) withthree ml. of petroleum ether. The petroleum ether The third group (the test group) was fed the diet with fraction was evaporated to dryness, and the resldue was the addition of 02% of z amino 4,6 diphenethylaminm 2 333222 122? g jgigz giz 3 2,: g a 2;: s-triazine and 2% of cholesterol in the basal diet. All rabp P yn bits were fed for 80 days, then sacrificed and examined.

and dlssolved m 2 of acetone' 10 Atherosclerotic lesions were macroscopically graded on a To this solution was added 4 ml. of a 1% solution of digitonin in aquenous ethanol and the mixture was scale of 0 to 5, according to the seventy of atheromatous left overnight to form the digitonide. The digitonide was Plaque- The results are shown In Table The Serum washed twice with 75% aqueous ethanol dissolved in 3 cholesterol level was not changed by the administration of ml. of pyridine and diluted to 10 ml. with dioxane. 2 ml. of 15 z'ammo 4,6 dlphenethylammo's'trlazmet but the the solution was added to 10 ml. of dioxane scintillator to P u ar edly decreased the cholesterol levels 1 n the estimate the activity. The results are shown in Table III; mam artery and m the hver- The macroscoplc examinatwn this Table clearly shows that 2-amino-4,6-diphenethylalso revealed that the title compound suppresses the deamino-s-triazine suppresses the bio-synthesis of cholesterol vclopment of arteriosclerosis from acetate in the liver.

TABLE Iv Choresterol Atheromata Cholesterol Weight in liver (graded 0-5) in main of liver (mgJg-m.) ------axtery(mg./gm.) (gm.) (mean-l-SD.) Arch. Thrac. Abdom. (mean+S.D.)

ssis 2. 50=|=0. 1s 0 0 0 1. :1:0. 2s 151=h14 23.49=l=2.65 4.4 3.3 2.0 19. 87=i;2. 47 1:105:20 =14. 29:1;3. 82 2.5 1.2 0.4 7.3812. 10

- Significant decrease (P 0.01) in comparison with the control group. Significant decrease (P 0.02) in comparison with the control group.

TABLE 111 What is claimed is: Proportion amino. Number 1. The method of treating arteriosclerosis in a human p -sof Total -P- 'l liver which comprises administering to a human in need of said in diet (percent) animals (mean'lsE') treatment from about 10 to 500 mg. doses of 2-amino-4,6-

References Cited I d.p.m." is an abridgement of "decay per minute."

Suter et al., Helvettca Ch1rn1ca Acta, vol. 48, 1965, pp. EXPERIMENT 40 1940 1944 Elfect of 2-amino-4,6-diphenethylamino-s-triazine on experimental arteriosclerosis in rabbits Three groups, each containing 5 rabbits, were used in JEROME D. GOLDBERG, Primary Examiner 

